Engineering

Publication Search Results

Now showing 1 - 10 of 16
  • (2022) Li, Yi
    Thesis
    CRISPR/Cas9-based gene editing is no doubt among the most intensively studied topics in bio-related fields in the recent decade, and certain new programmable Cas nucleases have been exploited recently for the development of a variety of biological tools far beyond gene editing, particularly for biosensor development. Although CRISPR/Cas-based biosensing has brought about a revolution in the area of nucleic acid diagnostic with their superior performances, its advantages were challenged when attempting to expand towards a broader range of non-nucleic acid targets. The currently reported methods for non-nucleic acid targets has successfully demonstrated the versality of CRISPR/Cas components in combining with other biosensing elements, however, combining these elements without proper optimization or controllable bioreaction environments could also bring additional variable factors into the system, hence potentially leading to compromised sensitivity or overall increase of system complexity. In this project, two novel CRISPR/Cas12a-based biosensing systems have been developed to realise simple, sensitive and universal non-nucleic acid detections. Both of these systems are established on a standard 96-well plate format, similar to the widely used ELISA diagnostic approach. The first system utilised an aptamer as its recognition molecule for rapid detection of two small proteins with fM-level sensitivity within 1.5 hours. In the second system, antibody was used as a recognition molecule, allowing to draw on a huge pool of commercially available antibodies to support its universality. This system exhibits ultra-high sensitivity down to 1 fg/mL (aM-level) for the detection of two protein targets. With these two successfully developed CRISPR/Cas12a-based non-nucleic acid biosensing systems, the universality and feasibility to deal with two practical bio-detection scenarios was investigated. The antibody-based system with minor modifications was directly used as a ready-to-use sensitivity enhancer onto a commercial IFN-γ ELISA kit, which resulted in a 2-log increase in sensitivity without changing its original protocol. Then, a similar modification strategy was used to re-direct the antibody-based system to detect whole pathogenic microorganism, Cryptosporidium parvum oocyst. Without the need for specialised instruments, the results show a successful detection of this pathogen with single oocyst sensitivity and capable of applying in challenging environmental samples. All these results serve as successful demonstration of the great potential in CRISPR/Cas-based biosensing technology to achieve affordable, translatable, and deployable solutions for various clinical, industrial and research diagnostic needs.

  • (2022) Cao, Jun
    Thesis
    This thesis focuses on the development and applications of magnetic resonance electrical properties tomography (MREPT), which is an emerging imaging modality to noninvasively obtain the electrical properties of tissues, such as conductivity and permittivity. Chapter 2 describes the general information about human research ethics, MRI scanner, MR sequence and the method of phase-based MREPT implemented in this thesis. Chapter 3 examines the repeatability of phase-based MREPT in the brain conductivity measurement using balanced fast field echo (bFFE) and turbo spin echo (TSE) sequences, and investigate the effects of compressed SENSE, whole-head B_1 shimming and video watching during scan on the measurement precision. Chapter 4 investigates the conductivity signal in response to short-duration visual stimulus, compares the signal and functional activation pathway with that of BOLD, and tests the consistency of functional conductivity imaging (funCI) with visual stimulation across participants. Chapter 5 extends the use of functional conductivity imaging to somatosensory stimulation and trigeminal nerve stimulation to evaluate the consistency of functional conductivity activation across different types of stimuli. In addition, visual adaptation experiment is performed to test if the repetition suppression effect can be observed using funCI. Chapter 6 explores if resting state conductivity networks can be reliably constructed using resting state funCI, evaluates the consistency of persistent homology architectures, and compares the links between nodes in the whole brain. Chapter 7 investigates the feasibility of prostate conductivity imaging using MREPT, and distinctive features in the conductivity distribution between healthy participants and participants with suspected abnormalities.

  • (2022) Qiao, Laicong
    Thesis
    There has been a rapid-growing market and academic enthusiasm for small wearable molecular diagnostic platforms driven by the growing demand for continuous monitoring of human health. Wearable devices need to be portable, stretchable, and ideally re-configurable to be able to work for different analytes. Such flexible physiological monitoring devices which are non-invasive or minimally-invasive represent the next frontier of biomedical diagnostics. They may make it possible to predict and prevent diseases or facilitate treatment by diagnosing diseases at the initial stages. However, there are many problems that restrict further applications of these devices. Firstly, there are a limited number of bio-materials which are highly flexible, biocompatible and have anti-fouling properties; such biomaterials are needed as substrates for wearable devices. Secondly, traditional biosensors used in wearable devices focus on the detection of physical signals (such as heartbeat) and small chemical molecules, e.g. Na+, K+. These are not sufficient to provide in depth health information which requires sensing of large molecules such as proteins, ideally in real time, which is currently challenging. This provides a motivation to develop highly sensitive wearable biosensors for the detection of large molecules in sweat. This thesis centres on the development of a bio-material based wearable device for continuous detection of crucial analytes in human sweat. To achieve this target, our first aim was to design a highly bio-compatible flexible material as a substrate for wearable devices. A tough and anti-fouling three-network hydrogel has been prepared by integrating a zwitterionic polymer network into a robust double-network hydrogel. Secondly, to fill the gap between technological development of continuous and non-invasive detection of different analytes in human sweat, a patterned sweat-based biosensor was created for the detection of key biomolecules. This sensor was produced by placing specific aptamers or enzymes on flexible working electrodes. In addition, nanotechnology methods have been applied to refine the bio-sensing interface to further increase the sensitivity of our sensors. Finally, a sample collection chip has been combined with our high sensitivity sensors to fabricate a wearable device for sweat bio-sensing purposes. Future research may involve integration of a commercially available wireless signal readout module with this wearable biosensing device. The outcomes of this work may provide new insights for the development of wearable devices for continuous measurement of a spectrum of analytes in sweat, as an important step towards point-of-care diagnostics

  • (2022) Luo, Jeff
    Thesis
    Nitric oxide (NO) plays pivotal roles in various physiological systems and has immense therapeutic potential. NO, however, has a short half-life (<5 s) and a short diffusion distance of ~160 μm in vivo, and its physiological functions are highly dependent on its concentrations. Current NO delivery strategies can be generally categorized into non-catalytic and catalytic (enzymatic) approaches. For the former, the longevity of the NO delivery systems principally relies on the finite NO donor reservoir, while the latter is limited by the low stability of natural enzymes. Another important challenge in NO delivery is the difficulty in accurately detecting circulating NO reservoir in blood. To address these challenges, this thesis focuses on the design, synthesis, and applications of nano-biomaterials to enable sustained NO delivery and accurate detection of endogenous circulating NO reservoir. This thesis revealed ceria nanoparticles as a new class of nanomaterials with the unique ability to catalyze NO generation from NO donors. The therapeutic activity of ceria-induced NO was demonstrated to inhibit cancer cell proliferation. This unique NO-generating feature stood in contrast to the well-established understanding of ceria to scavenge NO. This study provided deeper insights into the bio-functions of ceria nanoparticles and broadened their biomedical applications. Then this thesis reported the first catalytic polymers that generate NO, in particular amine-containing polymers, e.g., polyethyleneimine (PEI). These polymers can be easily integrated into a suite of biomaterials (e.g. hydrogels) to equip them with NO delivery capability. The therapeutic application of polymer-induced NO was demonstrated to prevent the formation of Pseudomonas aeruginosa biofilm. Finally, the thesis tackled the demand for rapid and accurate detection of human serum albumin (HSA, the most abundant circulating NO reservoir in blood) by developing a fluorescent paper-based sensor. This sensing platform allowed sensitive (detection limit of 0.91 g/L) and rapid (20 minutes) point-of-care detection of HSA and HSA-related disease diagnosis by visible color change, and could be extended to the detection of a spectrum of biomarkers. Collectively, these findings open new routes to produce next generation nano-biomaterials for the diverse biomedical applications of NO such as anticancer, antibacterial, and sensing applications.

  • (2022) Kaur, Jagjit
    Thesis
    Secreted by pancreatic β-cells, insulin is the major anabolic hormone, regulating the metabolism of fats, proteins, and carbohydrates. Defects in insulin production or action can lead to diabetes characterized by derangements in glucose handling and metabolic disease. Diabetes affects 420 million people worldwide, increasing morbidity, mortality and placing a burden on healthcare of nations. There is a need for rapid and accurate monitoring of insulin levels to optimize diabetes management and facilitate early diagnosis of insulin related chronic diseases. Conventional strategies such as HPLC, MALDI-TOF, ELISA, etc. used for insulin detection are not suitable for point-of-care testing (POCT) as they are expensive, and require sample preparation, sophisticated instruments, and skilled personnel. Our goal was to develop techniques to allow POCT for insulin in real time. In this study we developed two lateral flow assays (LFAs) based POCT platforms using aptamers as the biorecognition molecules for colorimetric and fluorescent detection of insulin. A range of conditions were tested such as concentrations of aptamers, reporter molecules used, volume of sample required, and assay time to obtain quantify insulin levels using a standard LFA reader. The colorimetric LFAs had linear detection range of 0.01-1 ng.mL-1 and LOD of 0.01 ng.mL-1. The fluorescent LFAs exhibited a linear detection range of 0-4 ng.mL-1 and 0.1 ng.mL-1 LOD. Various signal amplification strategies were incorporated, ie., gold-silver amplification technique and rolling circular amplification (RCA) to further enhance the signal. The developed colorimetric LFAs were successfully used for insulin quantification in rat blood, human blood, and human saliva samples. Although insulin levels were quantified within 12 min, some issues arose such as coagulation, need for dilution, and non-uniform flow through the test strips. Further work is required to optimize blood handling to progress an insulin POCT in real time. Future work could develop a multiplexed strip for detection of different analytes such as HbA1c, glucose, and C-peptide for better management of diabetes, along with a smartphone reader App. This research goes some way to addressing the challenge of providing a reliable and rapid approach for highly sensitive and specific detection of insulin for POCT applications.

  • (2006) Doran, Michael
    Thesis
    The transplantation of ex vivo expanded mobilized peripheral blood haematopoietic stem cells (PBSC), in place of unmanipulated cells following high dose chemotherapy, reduces the period of cytopenia associated with the therapy’s hemotoxicity. In this thesis the development and optimization of a preclinical prototype hollow fiber bioreactor (HFBR) for the ex vivo expansion of PBSC is described. Mass transport measurements and model of metabolite profiles demonstrate that Cuprophan and Polyflux are suitable membrane material for high-density cell expansion in a HFBR. Materials selected for the HFBR were found to be non-toxic following a 20-day saline extraction. Growth factor (GF) adsorption to the Polyflux membrane makes it unsuitable for expansion of GF dependent cells. However, the GF retention and minimal adsorption characteristics of the Cuprophan membrane are appropriate for this application. Cell-free medium degrades at 37ºC by an oxygen dependent process generating byproducts that inhibit cell growth. This process is relevant to perfusion bioreactors where the bulk of the medium is maintained at 37ºC and is cell-free. Albumin was shown to slow the degradation process but was itself degraded by shear damage inflicted during recirculation. Treating recirculating medium with dialysis against albumin was shown to be a more effective way to mitigate the effects of degradation and lengthen the functional life of albumin over conventional suspension of albumin in the recirculating medium. The preclinical prototype HFBR utilised dialysis against albumin to expand KG-1a cultures from densities as low as 3.5x10^5 cells/ml up to as high as 2x10^8 cells/ml with expansion rates equivalent to T-flask cultures. This process was then applied to PBSC where the targeted 100-fold expansion was achieved. Process optimization was continued using cord blood (CB) CD34+ cells. Growth factor loading sufficient for PBSC expansion in the HFBR was inadequate for CB expansions due to greater than anticipated CB uptake rates. The cell product from the HFBR contained significantly greater yields of CD34+ cells than attained using T-flask cultures. The HFBR platform is suitable for PBSC expansion and appears promising for CB expansion.

  • (2023) Vangelov, Belinda
    Thesis
    Background: Assessment of body composition, specifically evaluation of skeletal muscle (SM), has gained momentum in studies of patients with head and neck cancer (HNC). Depletion of SM measured via computed tomography (CT), known as CT-defined sarcopenia, has emerged as an independent prognostic indicator in HNC. International standard SM measures use the cross-sectional area (CSA) of a single axial slice at the third lumbar vertebra (L3). However, diagnostic CT scans for HNC do not always extend to this level, limiting assessment opportunities. This thesis investigates the feasibility of alternate vertebral levels for SM evaluation in HNC. Methods: A systematic review was undertaken to determine current evidence for SM evaluation at alternate vertebral levels in patients with cancer. Gaps in the literature led to a five phase plan to investigate the use of a cervical (C3) and thoracic (T2) level for SM assessment in patients with HNC who received a diagnostic or radiotherapy planning CT scan. This included evaluation of an existing prediction model (used to estimate L3-CSA with SM at C3), and formulation of population-specific models for use when L3 is not available. Novel methodology for SM evaluation at T2, and thresholds for low skeletal muscle index (SMI) values were also introduced. Results: The progressive findings of the five studies have indicated that; SM assessment at C3 should be applied with caution; prediction modelling should be population and sex-specific; thoracic SM measures at T2 deplete in similar proportions to L3 over time, cervical SM does not; SM at T2 is predictive of sarcopenia risk (HR=62.78, CI 27.59-164.08, p<0.001); and T2-SMI thresholds for sarcopenia stratified for sex and body mass index were effective in determining patients at risk of critical weight loss during treatments, and overall survival outcomes. Conclusion: This body of work has identified key concerns with the use of SM at C3 for muscle evaluation in patients with HNC, and has provided evidence for the use of SM at T2 as an alternative to L3. The anatomical position of T2 is not likely to include tumour infiltration, contains musculature that is sensitive to depletion, and is visible in CT scans taken in routine practice for HNC. Population and tumour-specific SMI thresholds for sarcopenia are required in this population for effective diagnosis and appropriate service delivery to ensure optimal nutritional and survival outcomes in this patient population.

  • (2023) Alduraywish, Abdulrahman
    Thesis
    Electroconvulsive therapy (ECT) is a neuromodulatory technique used widely for treatment of various psychiatric diseases such as Alzheimer’s and epilepsy. It includes the application of a large dose of electric current for short periods of time through attached electrodes on the scalp. However, there are some concerns related to electric field (EF) distribution in the brain and the potential for cognitive side effects. These concerns can be investigated thoroughly using computational modelling via the finite element (FE) method. To construct FE head models for simulating ECT stimulation, segmentation of magnetic resonance image (MRI) head scans is essential. This segmentation can be performed manually, which is laborious and time-consuming, or automatically. Various automatic segmentation approaches have been utilized to segment MRI head scans, including intensity-based techniques and deep learning techniques. In particular, convolutional neural networks (CNN), a subfield of deep learning, are considered as state-of-the-art owing to their outstanding performance. In this research, 2D U-Net and 3D U-Net CNN architectures were employed to rapidly segment seven MRI head scans into not only cerebrospinal fluid (CSF), grey matter (GM) and white matter (WM) as most studies performed but to include skin, skull tissues. This in turn will accelerate the process of construction of FE head models for simulating ECT. Two experiments were conducted using 2D U-Net, where in experiment 1, all the extracted slices from each head scan were used for training and testing, while experiment 2 involved only mid-slices. Additionally, two experiments were carried out using 3D U-Net with different patch sizes, 64 × 64 × 64 (experiment 3) and 128 × 128 × 128 (experiment 4). Inverse frequency class weighting was also used as an additional run for each experiment. The segmentation performance was evaluated in terms of intersection over union (IOU). In general, higher IOU scores were obtained for experiment 2 and experiment 4. The process of 3D segmentation (experiment 4) took only several seconds compared with previous approaches, which would take few hours. Predicted outputs from experiment 4 were used to construct FE models to simulate ECT. Various electrode montages were modelled on the scalp of each constructed head model, and EF magnitudes in various brain regions under ECT were compared against the corresponding manually-segmented head models. Larger EF magnitudes (10-25%) were exhibited in the automatically-segmented head models of four subjects, while weaker EFs (5-7%) were exhibited in the other three subjects compared with manual segmentation. These errors could be improved by increasing the training dataset to more than seven head scans, enhancing the segmentation accuracy for more accurate simulation of ECT.

  • (2023) Thai, Mai Thanh
    Thesis
    Gastrointestinal (GI) cancer is a leading cause of cancer death worldwide. Early removal of adenomatous polyps offers a fantastic outcome, with a 5-year survival rate exceeding 90%. Endoscopic submucosal dissection (ESD) is a minimally invasive approach to removing GI polyps, known for its high success in achieving en bloc resection. Despite the benefits of ESD with its low complication rates and short hospital stays, existing technologies have several limitations such as the need for multiple tools during the procedure and high force loss caused by rigid cables with nonlinear friction and backlash hysteresis. The nonlinearity can lead to imprecise control of surgical tools while the force loss hinders the tool's ability to access lesions located in the transverse and ascending colons. In addition, no endoscopic robots are equipped with soft 3D bioprinting capabilities to deliver in situ wound sealants for defect tissues. Finally, existing systems lack real-time force sensing and haptic display compared to open surgery, making optimal surgical procedures challenging. This thesis will develop a multifunctional and flexible soft robotic endoscopic system that can perform ESD and in situ 3D bioprinting. The new system features a high degree of freedom soft surgical head integrated into a long and flexible snake-like robotic arm that can access confined and hard-to-reach areas via small skin incisions or natural orifices. The device has a user-friendly design with a master-slave architecture and is operated by kinematic inversion models and a learning-based controller for precise tip motions. In addition, the system is equipped with a new 3D force sensor and an advanced haptic display that can efficiently reproduce the sense of touch for a better endoscopic procedure. The whole system is driven by a new soft microtubule artificial muscle (SMAM) that can be programmed to elongate and contract under hydraulic pressure. Mathematical models and advanced control algorithms are proposed to reduce the system's nonlinearity impact. The system's capability for endoscopic surgery with the haptic display is shown through two user studies and fresh porcine tissue. 3D printing abilities with various patterns on several surfaces and a colon phantom are further tested with different composite hydrogels and biomaterials. The new system is expected to fill a gap in the field of advanced endoscopic surgical robots while also assisting in the future growth of in situ bioprinting.

  • (2023) Yang, Biyao
    Thesis
    Triple negative breast cancer (TNBC) is a highly aggressive breast cancers with highest mortality rate and poorest outcomes. The challenges of TNBC treatment are rooted in lack of estrogen receptors (ER), progesterone receptors (PR), and human epidermal growth factor 2 (HER2), resulting endocrine therapy or receptors targeted therapies are not effective against TNBC. Clustered regularly interspaced short palindromic repeat-associated protein 9 (CRISPR/Cas9)-based gene therapy is a revolutionary strategy for TNBC treatment, while its clinical applications are limited by lack of safe and efficient non-viral delivery system. In this thesis, we developed light triggerable liposomes mediated CRISPR/Cas9 delivery for TNBC treatment. CRISPR/Cas9 was loaded inside liposomes with encapsulation efficiency higher than 70%. FDA-approved photosensitiser (PS) verteporfin was used to produce reactive oxygen species (ROS) when it was activated by external light, leading to liposome membrane destruction and thus release of CRISPR/Cas9 in cells. The knockdown of A20 gene-Exon3 by CRISPR/Cas9 in TNBC cells significantly supressed the cell proliferation and migration in vitro. CRISPR/Cas9-based gene editing via light triggerable liposomes with spatial and temporal control provides a promising option for TNBC treatment. The application of triggerable liposomes can be extended to deep tissue therapy by using X-ray as external stimuli. PS in liposomes not only works for liposome rupture, but also serves as photodynamic therapy (PDT) reagent to induce cell death through extensive ROS generation. X‑ray induced photodynamic therapy (X-PDT) via protoporphyrin IX (PPIX) & perfluorooctyl bromide (PFOB) co-loaded liposomes for in vitro TNBC treatment was developed. Relief of hypoxia in cancer cells by addition of PFOB improved ROS production with 4.9-fold of PPIX under 2Gy radiation and it turned out to induce 17% increase in cell death through either apoptosis or necrosis pathway. These findings in this thesis suggest that developed intelligent triggerable liposomes are superior to deliver CRISPR/Cas9 or photosensitisers efficiently for TNBC treatment. Future works were prospected to combine CRISPR/Cas9 gene editing with X-PDT via triggerable liposomes to achieve precise synthetic therapy.